Ado-resistant cysteamine analogs and uses thereof

ABSTRACT

The present disclosure is directed to methods for treating diseases for which cysteamine is indicated and compounds useful in such methods.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.15/201,969, which claims the benefit of U.S. Provisional Application No.62/187,939, filed Jul. 2, 2015, and U.S. Provisional Application No.62/387,337, filed Dec. 23, 2015, each of which is incorporated herein byreference in its entirety.

FIELD OF THE DISCLOSURE

The disclosure relates to materials and methods to treat diseases inwhich therapy with cysteamine is indicated. In particular, thedisclosure provides therapeutic methods involving administration to apatient of a compound as disclosed herein.

BACKGROUND

Cysteamine (HS-CH2-CH2-NH2) is a small sulfhydryl compound that is ableto cross cell membranes easily due to its small size. Cysteamine iscurrently FDA approved for use in the treatment of cystinosis, anintra-lysosomal cystine storage disorder. In cystinosis, cysteamine actsby converting cystine to cysteine and cysteine-cysteamine mixeddisulfide, which are then both able to leave the lysosome through thecysteine and lysine transporters respectively (Gahl et al., N Engl J Med347(2):111-21, 2002). Within the cytosol, the mixed disulfide can bereduced by its reaction with glutathione and the cysteine released canbe used for further GSH synthesis. Treatment with cysteamine has beenshown to result in lowering of intracellular cystine levels incirculating leukocytes (Dohil et al., J. Pediatr 148(6):764-9, 2006).

Cysteamine is converted to hypotaurine by cysteamine dioxygenase (ADO)(Coloso et al. (2006) Adv Exp Med Biol 583, 25-36; Dominy et al. (2007)J Biol Chem 282, 25189-25198; Richerson et al. (1987) Methods Enzymol143, 410-415) and then ultimately to taurine, the most common amino acidin the body. Cysteamine is also discussed in Prescott et al., (Lancet2(7778):652, 1979); Prescott et al., (Br Med J 1(6116):856-7, 1978);Mitchell et al., (Clin Pharmacol Ther 16(4):676-84, 1974); de Ferreyraet al., (Toxicol Appl Pharmacol. 48(2):221-8, 1979); and Qiu et al.,(World J Gastroenterol. 13:4328-32, 2007). Unfortunately, the sustainedconcentrations of cysteamine necessary for therapeutic effect aredifficult to maintain due to rapid metabolism and clearance ofcysteamine from the body, with nearly all administered cysteamineconverted to taurine in a matter of hours. These difficulties aretransferred to patients in the form of high dosing levels andfrequencies, with all of the consequent unpleasant side effectsassociated with cysteamine (e.g., gastrointestinal distress and bodyodor). See the package insert for CYSTAGON® (cysteamine bitartrate).International Publication No. WO 2007/079670 and U.S. Pat. Nos.8,026,2854 and 8,129,433 disclose enterically coated cysteamine productsand a method of reducing dosing frequency of cysteamine.

Cysteamine is addressed in International Patent Application Nos. WO2009/070781, and WO 2007/089670, and U.S. Patent Publication Nos.20110070272, 20090048154, and 20050245433.

SUMMARY

The present disclosure provides methods of treating a patient sufferingfrom a disease for which treatment with cysteamine is indicated. Themethods comprise administering to the patient an effective amount of acomposition comprising a compound as disclosed herein. It iscontemplated that administration of the composition reduces levels ofcystine in patients, which can improve the detrimental effects ofelevated cystine levels.

Suitable compositions comprise a compound of formula I or a disulfidethereof:

wherein:

R¹ and R² are independently selected from the group consisting of H andC₁₋₅alkyl; or

R¹ and R², taken together with the carbon atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring;

R³ and R⁴ are independently selected from the group consisting of H andC₁₋₅alkyl; or

R³ and R⁴, taken together with the carbon atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring;

G is selected from the group consisting of —NR⁵R⁶ and —CR⁷R⁸NR⁵R⁶;

R⁵ and R⁶ are independently selected from the group consisting of H andC₁₋₅alkyl; or

R⁵ and R⁶, taken together with the nitrogen atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered heterocyclic ring;

R⁷ and R⁸ are independently selected from the group consisting of H andC₁₋₅alkyl; or

R⁷ and R⁸, taken together with the carbon atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring;

R² and R⁶, taken together with the atoms to which they are attached,form a 4-, 5-, 6-, 7-, 8-, 9-, or 10-membered heterocyclic ring;

R⁴ and R⁶, taken together with the atoms to which they are attached,form a 4-, 5-, 6-, 7-, 8-, 9-, or 10-membered heterocyclic ring;

R² and R⁸, taken together with the atoms to which they are attached,form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring; or

R² and R⁴, taken together with the atoms to which they are attached,form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring.

In some cases, when G is —NH2, at least one of R¹, R², R³, and R⁴ isother than H.

In some cases, R⁵ and R⁶ are independently selected from the groupconsisting of H, methyl, and ethyl. In some cases, R⁵ and R⁶, takentogether with the nitrogen atom to which they are attached, form a5-membered heterocyclic ring.

In some cases, wherein R⁴ is methyl and/or R³ is methyl. In some cases,R³ and R⁴, taken together with the carbon atom to which they areattached, form a 3-membered carbocyclic ring.

In some cases, R² is methyl and/or R¹ is methyl. In some cases, R¹ andR², taken together with the carbon atom to which they are attached, forma 3-membered carbocyclic ring.

In some cases, G is —CR⁷R⁸NR⁵R⁶, and R² and R⁶, taken together with theatoms to which they are attached, form a 6-membered heterocyclic ring.In some cases, R⁵ is methyl.

In some cases, G is —NR⁵R⁶, and R² and R⁶, taken together with the atomsto which they are attached, form a 4- or 6-membered heterocyclic ring.In some cases, R⁵ is H.

In some cases, R⁷ and R⁸ are both H.

A compound of formula I includes, but is not limited to, the followingcompounds:

and disulfides thereof.

A compound of formula I includes, but is not limited to, the followingcompounds:

In some cases, R¹, R², R³, R⁴, R⁵, R⁶, R⁷, and R⁸ are independentlyselected from the group consisting of H and C₁₋₅alkyl. In some cases,R¹, R², R³, R⁴, R⁵, R⁶, R⁷, and R⁸ are independently selected from thegroup consisting of H and methyl.

Suitable compositions comprise a compound of formula II, formula III, ora disulfide thereof:

wherein:

L is a hydrocarbon linking group;

R⁹ and R¹⁰ are independently selected from the group consisting of H,C₁₋₅alkyl, and CO(C₁₋₅alkyl); or

R⁹ and R¹⁰, taken together with the nitrogen atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered heterocyclic ring;

A is a heterocyclic ring contain one N atom; and

n is 0, 1, 2, or 3.

In some cases, the compound of formula II is not cysteamine.

In some cases, the S atom in the compound of formula II or formula IIIis a distance of about 3.6 Angstroms to about 4.7 Angstroms from the Natom in the compound, such as about 3.8 Angstroms to about 4.4Angstroms, about 4.0 Angstroms to about 4.2 Angstroms, or about 4.1Angstroms from the N atom in the compound.

In some cases, L is a 3-, 4-, 5-, 6-, 7-, or 8-membered cycloalkyl ringor a 6-membered aryl ring. In some cases, L is C₁₋₅ alkyl. In somecases, L is substituted with one to four groups selected from halo, C₁₋₅alkyl, C₃₋₅ cycloalkyl, and —CO₂(C₁₋₅alkyl).

In some cases, A is a 3-, 4-, 5-, 6-, 7-, or 8-membered monocyclicheterocycloalkyl ring, a 6-, 7-, or 8-membered bicyclic heterocycloalkylring, or a 5- or 6-membered heteroaryl ring.

In some cases, the compound of formula III has a structure Ma:

wherein R¹¹ is selected from the group consisting of H and C₁₋₅ alkyl.

In some cases, A is substituted with one to four groups selected fromhalo, C₁₋₅alkyl, C₃₋₅ cycloalkyl, and —CO₂(C₁₋₅ alkyl).

In some cases, the compound of formula II, formula III, or disulfidethereof depletes cystine in a subject in an amount that is at least 70%of the level of depletion of cystine by cysteamine.

In various embodiments, the disclosure provides a compound as disclosedherein (e.g., a compound as represented by formula I, formula II, orformula III, or a disulfide thereof), wherein the compound or disulfidethereof produces reduced levels of dimethyl sulfide when administered toa subject compared to the level of dimethyl sulfide produced whencysteamine is administered to a subject. In some cases, least 2-foldless dimethyl sulfide is produced when the compound of formula I,formula II, or formula III, or disulfide thereof is administered to asubject.

In various embodiments, the disclosure provides a compound as disclosedherein (e.g., a compound as represented by formula I, formula II, orformula III, or a disulfide thereof), wherein the compound or disulfidethereof inhibits glutamate-induced excitotoxicity (i.e., providesneuroprotection). In some cases, the compound of formula I, formula II,or formula III, or disulfide thereof demonstrates at least 50% cellsurvival (expressed as a percent of the cell survival for 100 μMcysteamine), under conditions as described herein.

In various embodiments, the disclosure provides a method of treating apatient suffering from a disease for which treatment with cysteamine isindicated comprising administering to the patient an effective amount ofa composition comprising a compound as disclosed herein (e.g., acompound as represented by formula I, formula II, or formula III, or adisulfide thereof), wherein the compound or disulfide thereof isresistant to metabolism by cysteamine dioxygenase (ADO). In some cases,less than 20% of the compound of formula I, formula II, or formula III,or disulfide thereof is metabolized by ADO when assayed by consumptionof oxygen using an oxygen sensitive fluorescent probe.

Diseases for which treatment with cysteamine is indicated include, butare not limited to, cystinosis, fatty liver disease, fibrosis, athrombotic disease, an MECP-2 related disorder, an inheritedmitochondrial disease, a neurological disease or disorder, inflammationand cancer.

Fatty liver diseases include, but are not limited to, non-alcoholicfatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), fattyliver disease resulting from hepatitis, fatty liver disease resultingfrom obesity, fatty liver disease resulting from diabetes, fatty liverdisease resulting from insulin resistance, fatty liver disease resultingfrom hypertriglyceridemia, Abetalipoproteinemia, glycogen storagediseases, Weber-Christian disease, Wolmans disease, acute fatty liver ofpregnancy, and lipodystrophy.

Fibrosis includes, but is not limited to, atherosclerosis, asthma,cardiac fibrosis, organ transplant fibrosis, colloid and hypertrophicscar, muscle fibrosis, pancreatic fibrosis, bone-marrow fibrosis,interstitial liver fibrosis, cirrhosis of liver and gallbladder,scleroderma, pulmonary fibrosis, diffuse parenchymal lung disease,idiopathic interstitial fibrosis, interstitial pneumonitis, desquamativeinterstitial pneumonia, respiratory bronchiolitis, interstitial lungdisease, acute interstitial pneumonitis, nonspecific interstitialpneumonia, cryptogenic organizing pneumonia, lymphocytic interstitialpneumonia, renal fibrosis, and chronic kidney disease, cystic fibrosisand Alport's disease.

Thrombotic diseases include, but are not limited to, sickle celldisease, deep vein thrombosis, pulmonary embolism, cardiac embolism,hypercoagulable state, thrombophilia, Factor V Leiden, Antithrombin IIIdeficiency, Protein C deficiency, Protein S deficiency, Prothrombin genemutation (G20210A), Hyperhomcysteinemia, antiphospholipid antibodysyndrome (APS), anticardiolipin antibody (ACLA) thrombosis syndrome, orlupus anticoagulant (LA) syndrome.

Neurological diseases or disorders include, but are not limited to,Huntington's Disease, Parkinson's Disease, amyotrophic lateralsclerosis, multiple sclerosis, Alzheimer's disease spinal muscleatrophy, concussion, stroke, and traumatic brain injury (CTE).

MECP-2 related diseases include, but are not limited to, Rett syndrome,autism, pervasive development disorder, non-syndromic mentalretardation, idiopathic neonatal encephalopathy and idiopathic cerebralpalsy.

Inherited mitochondrial diseases include, but are not limited to,Friedreich's ataxia, Leber's hereditary optic neuropathy (LHON),myoclonic epilepsy and ragged-red fibers, mitochondrialencephalomyopathy, lactic acidosis, and stroke-like syndrome (MELAS),Kearn-Sayre syndrome and subacute necrotizing encephalopathy (Leigh'sSyndrome).

Cancers include, but are not limited to, breast cancer, melanoma,prostate cancer, pancreatic cancer, head and neck cancer, lung cancer,non small-cell lung carcinoma, renal cancer, colorectal cancer, coloncancer, ovarian cancer, liver cancer and gastric cancer.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph showing the relationship between the sulfur-nitrogendistance of a compound as disclosed herein compared to its level ofcystine depletion.

DETAILED DESCRIPTION Definitions

As used herein and in the appended claims, the singular forms “a,” “an,”and “the” include plural referents unless the context clearly dictatesotherwise. Thus, for example, reference to “a derivative” includes aplurality of such derivatives and reference to “a patient” includesreference to one or more patients and so forth.

Also, the use of “or” means “and/or” unless stated otherwise. Similarly,“comprise,” “comprises,” “comprising” “include,” “includes,” and“including” are interchangeable and not intended to be limiting.

It is to be further understood that where descriptions of variousembodiments use the term “comprising,” those skilled in the art wouldunderstand that in some specific instances, an embodiment can bealternatively described using language “consisting essentially of” or“consisting of.”

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood to one of ordinary skill inthe art to which this disclosure belongs. Although methods and materialssimilar or equivalent to those described herein can be used in thepractice of the disclosed methods and products, the exemplary methods,devices and materials are described herein.

The documents discussed above and throughout the text are providedsolely for their disclosure prior to the filing date of the presentapplication. Nothing herein is to be construed as an admission that theinventors are not entitled to antedate such disclosure by virtue ofprior disclosure. Each document is incorporated by reference in itsentirety with particular attention to the disclosure for which it iscited.

The following references provide one of skill with a general definitionof many of the terms used in this disclosure: Singleton, et al.,DICTIONARY OF MICROBIOLOGY AND MOLECULAR BIOLOGY (2d ed. 1994); THECAMBRIDGE DICTIONARY OF SCIENCE AND TECHNOLOGY (Walker ed., 1988); THEGLOSSARY OF GENETICS, 5TH ED., R. Rieger, et al. (eds.), Springer Verlag(1991); and Hale and Marham, THE HARPER COLLINS DICTIONARY OF BIOLOGY(1991).

As used herein, a “therapeutically effective amount” or “effectiveamount” refers to that amount of the compound sufficient to result inamelioration of symptoms, for example, treatment, healing, prevention oramelioration of the relevant medical condition, or an increase in rateof treatment, healing, prevention or amelioration of such conditions,typically providing a statistically significant improvement in thetreated patient population. When referencing an individual activeingredient, administered alone, a therapeutically effective dose refersto that ingredient alone. When referring to a combination, atherapeutically effective dose refers to combined amounts of the activeingredients that result in the therapeutic effect, whether administeredin combination, including serially or simultaneously. In someembodiments, such as for fatty liver disease, a therapeuticallyeffective amount of the compound ameliorates one or more symptoms,including but not limited to, liver fibrosis, fat content of liver,incidence of or progression of cirrhosis, incidence of hepatocellularcarcinoma, increased hepatic aminotransferase levels, such as ALT andAST, increased serum ferritin, elevated levels ofgamma-glutamyltransferase (gamma-GT), and elevated levels of plasmainsulin, cholesterol and triglyceride. In some embodiments, such as fora neurodegenerative disease, a therapeutically effective amount of thecompound increases the level of brain-derived neurotrophic factor(BDNF). In some embodiments, such as for a neurodegenerative disease, atherapeutically effective amount of the compound inhibits tissuetransglutaminase. In some embodiments, such as for a neurodegenerativedisease, a therapeutically effective amount of the compound increasesheat shock DnaJ-containing protein 1b.

As used herein “a disease for which treatment with cysteamine isindicated” refers to a disease in which increasing the level ofcysteamine and/or reducing the level of cystine in a patient isbeneficial. Contemplated diseases, include, but are not limited to anyof the diseases listed herein, including those in the “Indications,Dosing and Administration” section.

“Treatment” refers to prophylactic treatment or therapeutic treatment.In certain embodiments, “treatment” refers to administration of acompound or composition to a subject for therapeutic or prophylacticpurposes.

A “therapeutic” treatment is a treatment administered to a subject whoexhibits signs or symptoms of pathology for the purpose of diminishingor eliminating those signs or symptoms. The signs or symptoms may bebiochemical, cellular, histological, functional or physical, subjectiveor objective.

A “prophylactic” treatment is a treatment administered to a subject whodoes not exhibit signs of a disease or exhibits only early signs of thedisease, for the purpose of decreasing the risk of developing pathology.The compounds or compositions of the disclosure may be given as aprophylactic treatment to reduce the likelihood of developing apathology or to minimize the severity of the pathology, if developed.

“Diagnostic” means identifying the presence, extent and/or nature of apathologic condition. Diagnostic methods differ in their specificity andselectivity. While a particular diagnostic method may not provide adefinitive diagnosis of a condition, it suffices if the method providesa positive indication that aids in diagnosis.

“Pharmaceutical composition” refers to a composition suitable forpharmaceutical use in subject animal, including humans and mammals. Apharmaceutical composition comprises a therapeutically effective amountof a compound of the disclosure, optionally another biologically activeagent, and optionally a pharmaceutically acceptable excipient, carrieror diluent. In an embodiment, a pharmaceutical composition encompasses acomposition comprising the active ingredient(s), and the inertingredient(s) that make up the carrier, as well as any product thatresults, directly or indirectly, from combination, complexation oraggregation of any two or more of the ingredients, or from dissociationof one or more of the ingredients, or from other types of reactions orinteractions of one or more of the ingredients. Accordingly, thepharmaceutical compositions of the present disclosure encompass anycomposition made by admixing a compound of the disclosure and apharmaceutically acceptable excipient, carrier or diluent.

“Pharmaceutically acceptable carrier” refers to any of the standardpharmaceutical carriers, buffers, and the like, such as a phosphatebuffered saline solution, 5% aqueous solution of dextrose, and emulsions(e.g., an oil/water or water/oil emulsion). Non-limiting examples ofexcipients include adjuvants, binders, fillers, diluents, disintegrants,emulsifying agents, wetting agents, lubricants, glidants, sweeteningagents, flavoring agents, and coloring agents. Suitable pharmaceuticalcarriers, excipients and diluents are described in Remington'sPharmaceutical Sciences, 19th Ed. (Mack Publishing Co., Easton, 1995).Preferred pharmaceutical carriers depend upon the intended mode ofadministration of the active agent. Typical modes of administrationinclude enteral (e.g., oral) or parenteral (e.g., subcutaneous,intramuscular, intravenous or intraperitoneal injection; or topical,transdermal, or transmucosal administration).

A “pharmaceutically acceptable salt” is a salt that can be formulatedinto a compound for pharmaceutical use, including but not limited tometal salts (e.g., sodium, potassium, magnesium, calcium, etc.) andsalts of ammonia or organic amines.

As used herein “pharmaceutically acceptable” or “pharmacologicallyacceptable” is meant a material that is not biologically or otherwiseundesirable, i.e., the material may be administered to an individualwithout causing any undesirable biological effects or withoutinteracting in a deleterious manner with any of the components of thecomposition in which it is contained or with any components present onor in the body of the individual.

As used herein, the term “unit dosage form” refers to physicallydiscrete units suitable as unitary dosages for human and animalsubjects, each unit containing a predetermined quantity of a compound ofthe disclosure calculated in an amount sufficient to produce the desiredeffect, optionally in association with a pharmaceutically acceptableexcipient, diluent, carrier or vehicle. The specifications for the novelunit dosage forms of the present disclosure depend on the particularcompound employed and the effect to be achieved, and thepharmacodynamics associated with each compound in the host.

As used herein, the term “subject” encompasses mammals. Examples ofmammals include, but are not limited to, any member of the mammalianclass: humans, non-human primates such as chimpanzees, and other apesand monkey species; farm animals such as cattle, horses, sheep, goats,swine; domestic animals such as rabbits, dogs, and cats; laboratoryanimals including rodents, such as rats, mice and guinea pigs, and thelike. The term does not denote a particular age or gender. In variousembodiments the subject is human. In various embodiments, the subject isa child or adolescent.

In one aspect, a method is provided for treating a patient sufferingfrom a disease for which treatment with cysteamine is indicated. Themethod comprises administering to the patient an effective amount of acomposition comprising a compound of formula I or a disulfide thereof:

wherein:

R¹ and R² are independently selected from the group consisting of H andC₁₋₅alkyl; or

R¹ and R², taken together with the carbon atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring;

R³ and R⁴ are independently selected from the group consisting of H andC₁₋₅alkyl; or

R³ and R⁴, taken together with the carbon atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring;

G is selected from the group consisting of —NR⁵R⁶ and —CR⁷R⁸NR⁵R⁶;

R⁵ and R⁶ are independently selected from the group consisting of H andC₁₋₅alkyl; or

R⁵ and R⁶, taken together with the nitrogen atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered heterocyclic ring;

R⁷ and R⁸ are independently selected from the group consisting of H andC₁₋₅alkyl; or

R⁷ and R⁸, taken together with the carbon atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring;

R² and R⁶, taken together with the atoms to which they are attached,form a 4-, 5-, 6-, 7-, 8-, 9-, or 10-membered heterocyclic ring;

R⁴ and R⁶, taken together with the atoms to which they are attached,form a 4-, 5-, 6-, 7-, 8-, 9-, or 10-membered heterocyclic ring;

R² and R⁸, taken together with the atoms to which they are attached,form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring; or

R² and R⁴, taken together with the atoms to which they are attached,form a 3-, 4-, 5-, 6-, 7-, or 8-membered carbocyclic ring.

In some cases, when G is —NH2, at least one of R¹, R², R³, and R⁴ isother than H.

In some cases, R⁵ and R⁶ are independently selected from the groupconsisting of H, methyl, and ethyl. In some cases, R⁵ and R⁶, takentogether with the nitrogen atom to which they are attached, form a5-membered heterocyclic ring.

In some cases, wherein R⁴ is methyl and/or R³ is methyl. In some cases,R³ and R⁴, taken together with the carbon atom to which they areattached, form a 3-membered carbocyclic ring.

In some cases, R² is methyl and/or R¹ is methyl. In some cases, R¹ andR², taken together with the carbon atom to which they are attached, forma 3-membered carbocyclic ring.

In some cases, G is —CR⁷R⁸NR⁵R⁶, and R² and R⁶, taken together with theatoms to which they are attached, form a 6-membered heterocyclic ring.In some cases, R⁵ is methyl.

In some cases, G is —NR⁵R⁶, and R² and R⁶, taken together with the atomsto which they are attached, form a 4- or 6-membered heterocyclic ring.In some cases, R⁵ is H.

In some cases, R⁷ and R⁸ are both H.

A compound of formula I includes, but is not limited to, the followingcompounds:

and disulfides thereof.

A compound of formula I includes, but is not limited to, the followingcompounds:

In some cases, R¹, R², R³, R⁴, R⁵, R⁶, R⁷, and R⁸ are independentlyselected from the group consisting of H and C₁₋₅ alkyl. In some cases,R¹, R², R³, R⁴, R⁵, R⁶, R⁷, and R⁸ are independently selected from thegroup consisting of H and methyl.

In one aspect, a method is provided for treating a patient sufferingfrom a disease for which treatment with cysteamine is indicated. Themethod comprises administering to the patient an effective amount of acomposition comprising a compound of formula II, formula III, or adisulfide thereof:

wherein:

L is a hydrocarbon linking group;

R⁹ and R¹⁰ are independently selected from the group consisting of H,C₁₋₅ alkyl, and CO(C₁₋₅alkyl); or

R⁹ and R¹⁰, taken together with the nitrogen atom to which they areattached, form a 3-, 4-, 5-, 6-, 7-, or 8-membered heterocyclic ring;

A is a heterocyclic ring contain one N atom; and

n is 0, 1, 2, or 3.

In some cases, the compound of formula II is not cysteamine.

In some cases, the S atom in the compound of formula II or formula IIIis a distance of about 3.6 Angstroms to about 4.7 Angstroms from the Natom in the compound, such as about 3.8 Angstroms to about 4.4Angstroms, about 4.0 Angstroms to about 4.2 Angstroms, or about 4.1Angstroms from the N atom in the compound.

In some cases, L is a 3-, 4-, 5-, 6-, 7-, or 8-membered cycloalkyl ringor a 6-membered aryl ring. In some cases, L is C₁₋₅ alkyl. In somecases, L is substituted with one to four groups selected from halo, C₁₋₅alkyl, C₃₋₅ cycloalkyl, and —CO₂(C₁₋₅alkyl).

In some cases, A is a 3-, 4-, 5-, 6-, 7-, or 8-membered monocyclicheterocycloalkyl ring, a 6-, 7-, or 8-membered bicyclic heterocycloalkylring, or a 5- or 6-membered heteroaryl ring.

In some cases, the compound of formula III has a structure Ma:

wherein R¹¹ is selected from the group consisting of H and C₁₋₅ alkyl.

In some cases, A is substituted with one to four groups selected fromhalo, C₁₋₅alkyl, C₃₋₅ cycloalkyl, and —CO₂(C₁₋₅ alkyl).

Compounds disclosed herein also include, but are not limited to,

the compounds listed in the tables herein, and disulfides thereof.

Compounds disclosed herein include compounds having the followingstructure or a disulfide thereof:

wherein R¹ is C₁₋₅alkyl; and R³, R⁴, R⁵, R⁷, and R⁸ are independentlyselected from the group consisting of H and C₁₋₅alkyl.

Compounds disclosed herein include compounds having the followingstructure or a disulfide thereof:

wherein R¹, R³, R⁴, R⁵, R⁶, and R⁷ are independently selected from thegroup consisting of H and C₁₋₅ alkyl.

ADO-Resistant Cysteamine Analogs

The disclosure provides ADO-resistant cysteamine analogs for use in themethods described herein. An “ADO-resistant cysteamine analog” in thepresent disclosure refers generally to compounds of formula I, formulaII, formula III, or a disulfide thereof. ADO-resistant cysteamineanalogs generally demonstrate three properties: (1) the compounds areresistant to metabolism by ADO, (2) the compounds cleave cystine invivo, and (3) the compounds clear stored cystine in patient cystinoticfibroblasts.

As used herein a compound that is “resistant to metabolism by cysteaminedioxygenase” or “resistant to metabolism by ADO” refers to a compoundthat undergoes less than 50% degradation, for example, less than 40%,less than 30%, less than 25%, less than 20%, less than 15%, less than10%, less than 8%, less than 7%, less than 6%, less than 5%, less than4%, less than 3%, less than 2%, less than 1.5%, and/or less than 1%degradation when assayed in the presence of ADO under conditions asdescribed herein. Due to the rapid metabolism and clearance ofcysteamine from the body due to ADO, the sustained concentrations ofcysteamine necessary for therapeutic effect are difficult to maintain.Advantageously, compounds that are resistant to metabolism by ADO aremore readily maintained at necessary concentrations for therapeuticeffect.

A compound that cleaves cystine in vivo refers to a compound thatconverts cystine to cysteine and a mixed disulfide containing cysteineand the compound. Such compounds typically have a reactivity similar toor greater than the reactivity of cysteamine for cystine, such as atleast at least 50%, at least 75%, at least 90%, at least 100%, at least110%, at least 120%, at least 130%, at least 140%, at least 150%, atleast 160%, at least 170%, at least 180%, at least 190%, and/or at least200% of the reactivity of cysteamine for cystine, as determined underconditions as described herein.

A compound that clears stored cystine in patient cystinotic fibroblastsrefers to a compound that facilitates transport of cystine out oflysosomes. Such compounds typically deplete cystine in an amount similarto or greater than the depletion of cystine by cysteamine, such as atleast 25%, at least 40%, at least 50%, at least 60%, at least 70%, atleast 75%, at least 80%, at least 85%, at least 90%, at least 95%, atleast 100%, at least 105%, and/or at least 110% of the depletion bycysteamine, as determined under conditions as described herein.

The ADO-resistant cysteamine analogs also include biologically activemetabolites or derivatives thereof, and includes salts, esters, amides,alkylate compounds, prodrugs, analogs, phosphorylated compounds,sulfated compounds, or other chemically modified forms thereof (e.g.,chemically modified forms prepared by labeling with radionucleotides orenzymes and chemically modified forms prepared by attachment of polymerssuch as polyethylene glycol). Thus, the compounds of formula I, formulaII, or formula III can be administered in the form of apharmacologically acceptable salt, ester, amide, prodrug or analog or asa combination thereof.

Salts, esters, amides, prodrugs and analogs of the active agents may beprepared using standard procedures known to those skilled in the art ofsynthetic organic chemistry and described, for example, by J. March,“Advanced Organic Chemistry: Reactions, Mechanisms and Structure,” 4thEd. (New York: Wiley-Interscience, 1992). For example, basic additionsalts are prepared from the neutral drug using conventional means,involving reaction of one or more of the active agent's free hydroxylgroups with a suitable base. Generally, the neutral form of the drug isdissolved in a polar organic solvent such as methanol or ethanol and thebase is added thereto. The resulting salt either precipitates or may bebrought out of solution by addition of a less polar solvent. Suitablebases for forming basic addition salts include, but are not limited to,inorganic bases such as sodium hydroxide, potassium hydroxide, ammoniumhydroxide, calcium hydroxide, trimethylamine, or the like. Preparationof esters involves functionalization of hydroxyl groups which may bepresent within the molecular structure of the drug. The esters aretypically acyl-substituted derivatives of free alcohol groups, i.e.,moieties which are derived from carboxylic acids of the formula R—COOHwhere R is alkyl, and typically is lower alkyl. Esters can bereconverted to the free acids, if desired, by using conventionalhydrogenolysis or hydrolysis procedures. Preparation of amides andprodrugs can be carried out in an analogous manner. Other derivativesand analogs of the active agents may be prepared using standardtechniques known to those skilled in the art of synthetic organicchemistry, or may be deduced by reference to the pertinent literature.

Dimethyl Sulfide (DMS) Production

Compounds that produce reduced levels of dimethyl sulfide whenadministered to a subject (compared to the level produced whencysteamine is administered) are desirable because unpleasant sideeffects associated with cysteamine (e.g., halitosis) may be reduced. Acompound that produces reduced levels of dimethyl sulfide whenadministered to a subject generally produces at least 2-fold lessdimethyl sulfide, such as at least 3-fold less, 4-fold less, 5-foldless, 6-fold less, 8-fold less, 10-fold less, 15-fold less, and/or20-fold less dimethyl sulfide, compared to the level of dimethyl sulfideproduced when cysteamine is administered to the subject at the same doseand same time after administration under conditions as described herein.For example, the level of dimethyl sulfide can be measured byadministering the compound in a dose of about 10 mg/kg to about 500mg/kg, such as about 25 mg/kg to about 400 mg/kg, about 50 mg/kg toabout 300 mg/kg, about 75 mg/kg to about 200 mg/kg, about 10 mg/kg,about 25 mg/kg, about 50 mg/kg, about 75 mg/kg, about 100 mg/kg, about150 mg/kg, about 200 mg/kg, about 300 mg/kg, about 400 mg/kg, and/orabout 500 mg/kg, and by measuring the dimethyl sulfide level about 10minutes to 4 hours after administration, such as about 15 minutes toabout 2 hours, about 30 minutes to about 1 hour, about 15 minutes, about30 minutes, about 1 hour, and/or about 2 hours after administration.

Neuroprotection

Excitotoxicity disorders affect the central nervous and peripheralnervous systems and can lead to progressive neurodegeneration.Excitotoxicity results from excess glutamate being secreted by variouscells, including immune cells and neurons, in the brain. Glutamate isthe primary excitatory neurotransmitter in the mammalian nervous system.Prolonged glutamate signaling leads to a type of toxicity characterizedby elevated mitochondrial activity, gradual glutathione (GSH) depletion,oxidative stress and apoptosis (Shih et al., J Neurosci. 26:10514-523,2006). Cysteamine is capable of inhibiting glutamate-inducedexcitotoxicity in St-HdhQ^(111/111) cells. A compound that inhibitsglutamate-induced excitotoxicity (i.e., provides neuroprotection)generally provides at least 50% cell survival (expressed as a percent ofthe cell survival for 100 μM cysteamine), such as at least 65%, at least75%, at least 80%, at least 90%, and/or at least 95% cell survival,under conditions as described herein.

Pharmaceutical Formulations

The disclosure provides compounds useful in the treatment of diseases inwhich therapy with cysteamine is indicated. To administer compounds ofthe disclosure to patients or test animals, it is preferable toformulate the compounds in a composition comprising one or morepharmaceutically acceptable carriers. Pharmaceutically orpharmacologically acceptable carriers or vehicles refer to molecularentities and compositions that do not produce allergic, or other adversereactions when administered using routes well-known in the art, asdescribed below, or are approved by the U.S. Food and DrugAdministration or a counterpart foreign regulatory authority as anacceptable additive to orally or parenterally administeredpharmaceuticals. Pharmaceutically acceptable carriers include any andall clinically useful solvents, dispersion media, coatings,antibacterial and antifungal agents, isotonic and absorption delayingagents and the like.

Pharmaceutical carriers include pharmaceutically acceptable salts,particularly where a basic or acidic group is present in a compound. Forexample, when an acidic substituent, such as —COOH, is present, theammonium, sodium, potassium, calcium and the like salts, arecontemplated for administration. Additionally, where an acid group ispresent, pharmaceutically acceptable esters of the compound (e.g.,methyl, tert-butyl, pivaloyloxymethyl, succinyl, and the like) arecontemplated as preferred forms of the compounds, such esters beingknown in the art for modifying solubility and/or hydrolysischaracteristics for use as sustained release or prodrug formulations.

When a basic group (such as amino or a basic heteroaryl radical, such aspyridyl) is present, then an acidic salt, such as hydrochloride,hydrobromide, acetate, maleate, pamoate, phosphate, methanesulfonate,p-toluenesulfonate, and the like, is contemplated as a form foradministration.

In addition, compounds may form solvates with water or common organicsolvents. Such solvates are contemplated as well.

The compounds may be administered orally, parenterally, transocularly,intranasally, transdermally, transmucosally, by inhalation spray,vaginally, rectally, or by intracranial injection. The term parenteralas used herein includes subcutaneous injections, intravenous,intramuscular, intracisternal injection, or infusion techniques.Administration by intravenous, intradermal, intramusclar, intramammary,intraperitoneal, intrathecal, retrobulbar, intrapulmonary injection andor surgical implantation at a particular site is contemplated as well.Generally, compositions for administration by any of the above methodsare essentially free of pyrogens, as well as other impurities that couldbe harmful to the recipient. Further, compositions for administrationparenterally are sterile.

Pharmaceutical compositions of the disclosure containing a compound asdisclosed herein as an active ingredient may contain pharmaceuticallyacceptable carriers or additives depending on the route ofadministration. Examples of such carriers or additives include water, apharmaceutically acceptable organic solvent, collagen, polyvinylalcohol, polyvinylpyrrolidone, a carboxyvinyl polymer,carboxymethylcellulose sodium, polyacrylic sodium, sodium alginate,water-soluble dextran, carboxymethyl starch sodium, pectin, methylcellulose, ethyl cellulose, xanthan gum, gum Arabic, casein, gelatin,agar, diglycerin, glycerin, propylene glycol, polyethylene glycol,Vaseline, paraffin, stearyl alcohol, stearic acid, human serum albumin(HSA), mannitol, sorbitol, lactose, a pharmaceutically acceptablesurfactant and the like. Additives used are chosen from, but not limitedto, the above or combinations thereof, as appropriate, depending on thedosage form of the disclosure.

Formulation of the pharmaceutical composition will vary according to theroute of administration selected (e.g., solution, emulsion). Anappropriate composition comprising the compound to be administered canbe prepared in a physiologically acceptable vehicle or carrier. Forsolutions or emulsions, suitable carriers include, for example, aqueousor alcoholic/aqueous solutions, emulsions or suspensions, includingsaline and buffered media. Parenteral vehicles can include sodiumchloride solution, Ringer's dextrose, dextrose and sodium chloride,lactated Ringer's or fixed oils. Intravenous vehicles can includevarious additives, preservatives, or fluid, nutrient or electrolytereplenishers.

A variety of aqueous carriers, e.g., water, buffered water, 0.4% saline,0.3% glycine, or aqueous suspensions may contain the active compound inadmixture with excipients suitable for the manufacture of aqueoussuspensions. Such excipients are suspending agents, for example sodiumcarboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose,sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia;dispersing or wetting agents may be a naturally-occurring phosphatide,for example lecithin, or condensation products of an alkylene oxide withfatty acids, for example polyoxyethylene stearate, or condensationproducts of ethylene oxide with long chain aliphatic alcohols, forexample heptadecaethyleneoxycetanol, or condensation products ofethylene oxide with partial esters derived from fatty acids and ahexitol such as polyoxyethylene sorbitol monooleate, or condensationproducts of ethylene oxide with partial esters derived from fatty acidsand hexitol anhydrides, for example polyethylene sorbitan monooleate.The aqueous suspensions may also contain one or more preservatives, forexample ethyl, or n-propyl, p-hydroxybenzoate, one or more coloringagents, one or more flavoring agents, and one or more sweetening agents,such as sucrose or saccharin.

In some embodiments, the compounds of this disclosure can be lyophilizedfor storage and reconstituted in a suitable carrier prior to use. Anysuitable lyophilization and reconstitution techniques can be employed.It is appreciated by those skilled in the art that lyophilization andreconstitution can lead to varying degrees of activity loss and that uselevels may have to be adjusted to compensate.

Dispersible powders and granules suitable for preparation of an aqueoussuspension by the addition of water provide the active compound inadmixture with a dispersing or wetting agent, suspending agent and oneor more preservatives. Suitable dispersing or wetting agents andsuspending agents are exemplified by those already mentioned above.Additional excipients, for example sweetening, flavoring and coloringagents, may also be present.

In one embodiment, the disclosure provides use of an enterically coatedcomposition. Enteric coatings prolong release until the active agentreaches the intestinal tract, typically the small intestine. Because ofthe enteric coatings, delivery to the small intestine is improvedthereby improving uptake of the active ingredient while reducing gastricside effects. Exemplary enterically coated products are described inInternational Publication No. WO 2007/089670 published Aug. 9, 2007,which is incorporated in its entirety herein.

In some embodiments, the coating material is selected such that thetherapeutically active agent is released when the dosage form reachesthe small intestine or a region in which the pH is greater than pH 4.5.The coating may be a pH-sensitive materials, which remain intact in thelower pH environs of the stomach, but which disintegrate or dissolve atthe pH commonly found in the small intestine of the patient. Forexample, the enteric coating material begins to dissolve in an aqueoussolution at pH between about 4.5 to about 5.5 or between about 5.5 to6.5. For example, pH-sensitive materials will not undergo significantdissolution until the dosage form has emptied from the stomach. The pHof the small intestine gradually increases from about 4.5 to about 6.5in the duodenal bulb to about 7.2 in the distal portions of the smallintestine. In order to provide predictable dissolution corresponding tothe small intestine transit time of about 3 hours (e.g., 2-3 hours) andpermit reproducible release therein, the coating should begin todissolve at the pH range within the small intestine. Therefore, theamount of enteric polymer coating should be sufficient to substantiallydissolve during the approximate three hour transit time within the smallintestine, such as the proximal and mid-intestine.

Enteric coatings have been used for many years to arrest the release ofthe drug from orally ingestible dosage forms. Depending upon thecomposition and/or thickness, the enteric coatings are resistant tostomach acid for required periods of time before they begin todisintegrate and permit release of the drug in the lower stomach orupper part of the small intestines. Examples of some enteric coatingsare disclosed in U.S. Pat. No. 5,225,202 which is incorporated byreference fully herein. As set forth in U.S. Pat. No. 5,225,202, someexamples of coating previously employed are beeswax and glycerylmonostearate; beeswax, shellac and cellulose; and cetyl alcohol, masticand shellac, as well as shellac and stearic acid (U.S. Pat. No.2,809,918); polyvinyl acetate and ethyl cellulose (U.S. Pat. No.3,835,221); and neutral copolymer of polymethacrylic acid esters(Eudragit L30D) (F. W. Goodhart et al., Pharm. Tech., pp. 64-71, April1984); copolymers of methacrylic acid and methacrylic acid methylester(Eudragits), or a neutral copolymer of polymethacrylic acid esterscontaining metallic stearates (Mehta et al., U.S. Pat. Nos. 4,728,512and 4,794,001). Such coatings comprise mixtures of fats and fatty acids,shellac and shellac derivatives and the cellulose acid phthlates, e.g.,those having a free carboxyl content. See, Remington's at page 1590, andZeitova et al. (U.S. Pat. No. 4,432,966), for descriptions of suitableenteric coating compositions. Accordingly, increased adsorption in thesmall intestine due to enteric coatings of product compositions canresult in improved efficacy.

Generally, the enteric coating comprises a polymeric material thatprevents product release in the low pH environment of the stomach butthat ionizes at a slightly higher pH, typically a pH of 4 or 5, and thusdissolves sufficiently in the small intestines to gradually release theactive agent therein. Accordingly, among the most effective entericcoating materials are polyacids having a pKa in the range of about 3 to5. Suitable enteric coating materials include, but are not limited to,polymerized gelatin, shellac, methacrylic acid copolymer type CNF,cellulose butyrate phthalate, cellulose hydrogen phthalate, celluloseproprionate phthalate, polyvinyl acetate phthalate (PVAP), celluloseacetate phthalate (CAP), cellulose acetate trimellitate (CAT),hydroxypropyl methylcellulose phthalate, hydroxypropyl methylcelluloseacetate, dioxypropyl methylcellulose succinate, carboxymethylethylcellulose (CMEC), hydroxypropyl methylcellulose acetate succinate(HPMCAS), and acrylic acid polymers and copolymers, typically formedfrom methyl acrylate, ethyl acrylate, methyl methacrylate and/or ethylmethacrylate with copolymers of acrylic and methacrylic acid esters(Eudragit NE, Eudragit RL, Eudragit RS). In one embodiment, the productcomposition is administered in oral delivery vehicle, including but notlimited to, tablet or capsule form. Tablets are manufactured by firstenterically coating the product. A method for forming tablets herein isby direct compression of the powders containing the enterically coatedproduct, optionally in combination with diluents, binders, lubricants,disintegrants, colorants, stabilizers or the like. As an alternative todirect compression, compressed tablets can be prepared usingwet-granulation or dry-granulation processes. Tablets may also be moldedrather than compressed, starting with a moist material containing asuitable water-soluble lubricant.

In a further embodiment, the product is formulated as a capsule. In oneembodiment, the capsule comprises the product and the capsule is thenenterically coated. Capsule formulations are prepared using techniquesknown in the art.

The preparation of delayed, controlled or sustained/extended releaseforms of pharmaceutical compositions with the desired pharmacokineticcharacteristics is known in the art and can be accomplished by a varietyof methods. For example, oral controlled delivery systems includedissolution-controlled release (e.g., encapsulation dissolution controlor matrix dissolution control), diffusion-controlled release (reservoirdevices or matrix devices), ion exchange resins, osmotic controlledrelease or gastroretentive systems. Dissolution controlled release canbe obtained, e.g., by slowing the dissolution rate of a drug in thegastrointestinal tract, incorporating the drug in an insoluble polymer,and coating drug particles or granules with polymeric materials ofvarying thickness. Diffusion controlled release can be obtained, e.g.,by controlling diffusion through a polymeric membrane or a polymericmatrix. Osmotically controlled release can be obtained, e.g., bycontrolling solvent influx across a semipermeable membrane, which inturn carries the drug outside through a laser-drilled orifice. Theosmotic and hydrostatic pressure differences on either side of themembrane govern fluid transport. Prolonged gastric retention may beachieved by, e.g., altering density of the formulations, bioadhesion tothe stomach lining, or increasing floating time in the stomach. Forfurther detail, see the Handbook of Pharmaceutical Controlled ReleaseTechnology, Wise, ed., Marcel Dekker, Inc., New York, N.Y. (2000),incorporated by reference herein in its entirety, e.g. Chapter 22 (“AnOverview of Controlled Release Systems”).

The concentration of product in these formulations can vary widely, forexample from less than about 0.5%, usually at or at least about 1% to asmuch as 15 or 20% by weight and are selected primarily based on fluidvolumes, manufacturing characteristics, viscosities, etc., in accordancewith the particular mode of administration selected. Actual methods forpreparing administrable compositions are known or apparent to thoseskilled in the art and are described in more detail in, for example,Remington's Pharmaceutical Science, 15th ed., Mack Publishing Company,Easton, Pa. (1980).

Compositions useful for administration may be formulated with uptake orabsorption enhancers to increase their efficacy. Such enhancers include,for example, salicylate, glycocholate/linoleate, glycholate, aprotinin,bacitracin, SDS, caprate and the like. See, e.g., Fix (J. Pharm. Sci.,85:1282-1285, 1996) and Oliyai and Stella (Ann. Rev. Pharmacol.Toxicol., 32:521-544, 1993).

The enterically coated product can comprise various excipients, as iswell known in the pharmaceutical art, provided such excipients do notexhibit a destabilizing effect on any components in the composition.Thus, excipients such as binders, bulking agents, diluents,disintegrants, lubricants, fillers, carriers, and the like can becombined with the cysteamine product. Oral delivery vehiclescontemplated for use herein include tablets or capsules. For solidcompositions, diluents are typically necessary to increase the bulk of atablet or capsule so that a practical size is provided for compression.Suitable diluents include dicalcium phosphate, calcium sulfate, lactose,cellulose, kaolin, mannitol, sodium chloride, dry starch and powderedsugar. Binders are used to impart cohesive qualities to a oral deliveryvehicle formulation, and thus ensure that a tablet remains intact aftercompression. Suitable binder materials include, but are not limited to,starch (including corn starch and pregelatinized starch), gelatin,sugars (including sucrose, glucose, dextrose and lactose), polyethyleneglycol, waxes, and natural and synthetic gums, e.g., acacia sodiumalginate, polyvinylpyrrolidone, cellulosic polymers (includinghydroxypropyl cellulose, hydroxypropyl methylcellulose, methylcellulose, hydroxyethyl cellulose, hypromellose, and the like), andVeegum. Lubricants are used to facilitate oral delivery vehiclemanufacture; examples of suitable lubricants include, for example,magnesium stearate, calcium stearate, and stearic acid, and aretypically present at no more than approximately 1 weight percentrelative to tablet weight. Disintegrants are used to facilitate oraldelivery vehicle, (e.g., a tablet) disintegration or “breakup” afteradministration, and are generally starches, clays, celluloses, algins,gums or crosslinked polymers. If desired, the pharmaceutical compositionto be administered may also contain minor amounts of nontoxic auxiliarysubstances such as wetting or emulsifying agents, pH buffering agentsand the like, for example, sodium acetate, sorbitan monolaurate,triethanolamine sodium acetate, triethanolamine oleate, and the like. Ifdesired, flavoring, coloring and/or sweetening agents may be added aswell. Other optional components for incorporation into an oralformulation herein include, but are not limited to, preservatives,suspending agents, thickening agents, and the like. Fillers include, forexample, insoluble materials such as silicon dioxide, titanium oxide,alumina, talc, kaolin, powdered cellulose, microcrystalline cellulose,and the like, as well as soluble materials such as mannitol, urea,sucrose, lactose, dextrose, sodium chloride, sorbitol, and the like.

A pharmaceutical composition may also comprise a stabilizing agent suchas hydroxypropyl methylcellulose or polyvinylpyrrolidone, as disclosedin U.S. Pat. No. 4,301,146. Other stabilizing agents include, but arenot limited to, cellulosic polymers such as hydroxypropyl cellulose,hydroxyethyl cellulose, methyl cellulose, ethyl cellulose, celluloseacetate, cellulose acetate phthalate, cellulose acetate trimellitate,hydroxypropyl methylcellulose phthalate, microcrystalline cellulose andcarboxymethylcellulose sodium; and vinyl polymers and copolymers such aspolyvinyl acetate, polyvinylacetate phthalate, vinylacetate crotonicacid copolymer, and ethylene-vinyl acetate copolymers. The stabilizingagent is present in an amount effective to provide the desiredstabilizing effect; generally, this means that the ratio of cysteamineproduct to the stabilizing agent is at least about 1:500 w/w, morecommonly about 1:99 w/w.

The tablet, capsule or other oral delivery system is manufactured byfirst enterically coating the product. A method for forming tabletsherein is by direct compression of the powders containing theenterically coated cysteamine product, optionally in combination withdiluents, binders, lubricants, disintegrants, colorants, stabilizers orthe like. As an alternative to direct compression, compressed tabletscan be prepared using wet-granulation or dry-granulation processes.Tablets may also be molded rather than compressed, starting with a moistmaterial containing a suitable water-soluble lubricant.

In an alternative embodiment, the enterically coated product isgranulated and the granulation is compressed into a tablet or filledinto a capsule. Capsule materials may be either hard or soft, and aretypically sealed, such as with gelatin bands or the like. Tablets andcapsules for oral use will generally include one or more commonly usedexcipients as discussed herein.

For administration of the dosage form, i.e., the tablet or capsulecomprising the enterically coated product, a total weight in the rangeof approximately 100 mg to 1000 mg is used. The dosage form is orallyadministered to a patient suffering from a condition for which treatmentwith cysteamine would typically be indicated.

Indications, Dosing and Administration

The disclosure provides methods of treating a patient (e.g., a humanpatient) suffering from a disease where therapy with cysteamine isindicated, comprising administering to the patient a therapeuticallyeffective amount of a compound as disclosed herein.

In some embodiments of the methods, the disease is cystinosis. In someembodiments the disease is nephropathic cystinosis. In some embodimentsof the methods, the disease is a fatty liver disease. In someembodiments, the fatty liver disease is non-alcoholic fatty liverdisease (NAFLD), non-alcoholic steatohepatitis (NASH), fatty liverdisease resulting from hepatitis, fatty liver disease resulting fromobesity, fatty liver disease resulting from diabetes, fatty liverdisease resulting from insulin resistance, fatty liver disease resultingfrom hypertriglyceridemia, Abetalipoproteinemia, glycogen storagediseases, Weber-Christian disease, Wolmans disease, acute fatty liver ofpregnancy, and lipodystrophy or other fatty liver disease. The term“fatty liver disease” may include or exclude NASH. In some embodimentsof the methods, the disease is a fibrosis. In some embodiments, thefibrosis is atherosclerosis, asthma, cardiac fibrosis, organ transplantfibrosis, colloid and hypertrophic scar, muscle fibrosis, pancreaticfibrosis, bone-marrow fibrosis, interstitial liver fibrosis, cirrhosisof liver and gallbladder, scleroderma, pulmonary fibrosis, diffuseparenchymal lung disease, idiopathic interstitial fibrosis, interstitialpneumonitis, desquamative interstitial pneumonia, respiratorybronchiolitis, interstitial lung disease, acute interstitialpneumonitis, nonspecific interstitial pneumonia, cryptogenic organizingpneumonia, lymphocytic interstitial pneumonia, renal fibrosis, chronickidney disease, cystic fibrosis, or Alport's disease. In someembodiments of the methods, the disease is a thrombotic disease. In someembodiments, the thrombotic disease is sickle cell disease, deep veinthrombosis, pulmonary embolism, cardiac embolism, hypercoagulable state,thrombophilia, Factor V Leiden, Antithrombin III deficiency, Protein Cdeficiency, Protein S deficiency, Prothrombin gene mutation (G20210A),Hyperhomcysteinemia, antiphospholipid antibody syndrome (APS),anticardiolipin antibody (ACLA) thrombosis syndrome, or lupusanticoagulant (LA) syndrome. In some embodiments of the methods, thedisease is an MECP-2 related disorder such as Rett syndrome, autism,pervasive development disorder, non-syndromic mental retardation,idiopathic neonatal encephalopathy or idiopathic cerebral palsy. In someembodiments of the methods, the disease is an inherited mitochondrialdisease such as Friedreich's ataxia, Leber's hereditary optic neuropathy(LHON), myoclonic epilepsy and ragged-red fibers, mitochondrialencephalomyopathy, lactic acidosis, and stroke-like syndrome (MELAS),Kearn-Sayre syndrome or subacute necrotizing encephalopathy (Leigh'sSyndrome). In some embodiments of the methods, the disease is aneurological disease or disorder such as Huntington's Disease,Parkinson's Disease, amyotrophic lateral sclerosis, multiple sclerosis,Alzheimer's disease spinal muscle atrophy, concussion, stroke, ortraumatic brain injury (CTE). In some embodiments of the methods, thedisease is inflammation. In some embodiments of the methods, the diseaseis cancer, for example, breast cancer, melanoma, prostate cancer,pancreatic cancer, head and neck cancer, lung cancer, non small-celllung carcinoma, renal cancer, colorectal cancer, colon cancer, ovariancancer, liver cancer or gastric cancer.

As used herein, “renal fibrosis or chronic kidney disease” refers to aprogressive disorder of the kidney characterized by excessive deposit(s)of extracellular matrix (ECM) and resulting in glomerular sclerosis andrenal tubule-interstitium fibrosis. Excessive deposit of fibrous tissuereplaces healthy kidney tissue, damaging kidney structure and impairingkidney function. Exemplary renal fibrosis or chronic kidney diseaseinclude, but are not limited to, chronic renal insufficiency (CRI),stage III, IV or V chronic kidney disease, nephropathy,glomerulosclerosis, glomerulonephritis, diabetes, fibrocystic kidneydisease, fibrotic kidney cancer, or renal interstitial fibrosis.

The compound is administered in a therapeutically effective amount. Theamount of compound to be administered is dependent on the age, weight,and general condition of the patient, the severity of the conditionbeing treated, and the judgment of the prescribing-physician. Suitabletherapeutic amounts are determined by standard methods by those skilledin the art. In some embodiments, the dose is administered either one ortwo times per day. In some embodiments, the dose is administeredmultiple times per day. In some embodiments, an effective dosage ofcompound is within the range of about 0.01 mg to about 1000 mg per kg(mg/kg) of body weight per day. Further, the effective dose may beabout: 0.5 mg/kg, 1 mg/kg, 5 mg/kg, 10 mg/kg, 15 mg/kg, 20 mg/kg/25mg/kg, 30 mg/kg, 35 mg/kg, 40 mg/kg, 45 mg/kg, 50 mg/kg, 55 mg/kg, 60mg/kg, 70 mg/kg, 75 mg/kg, 80 mg/kg, 90 mg/kg, 100 mg/kg, 125 mg/kg, 150mg/kg, 175 mg/kg, 200 mg/kg, and may increase by 25 mg/kg increments upto 1000 mg/kg, or may range between any two of the foregoing values. Insome embodiments, the administration route is oral. In some embodiments,the administration route is transdermal.

The compound is administered in a therapeutically effective amount;typically, the composition is in unit dosage form. The amount ofcompound administered is, of course, dependent on the age, weight, andgeneral condition of the patient, the severity of the condition beingtreated, and the judgment of the prescribing physician. Suitabletherapeutic amounts will be known to those skilled in the art and/or aredescribed in the pertinent reference texts and literature. In oneaspect, the dose is administered either one time per day or multipletimes per day. The product may be administered one, two or three or fourtimes per day. In some embodiments, an effective dosage of product maybe within the range of 0.01 mg to 1000 mg per kg (mg/kg) of body weightper day. Further, the effective dose may be 0.5 mg/kg, 1 mg/kg, 5 mg/kg,10 mg/kg, 15 mg/kg, 20 mg/kg, 25 mg/kg, 30 mg/kg, 35 mg/kg, 40 mg/kg, 45mg/kg, 50 mg/kg, 55 mg/kg, 60 mg/kg, 70 mg/kg, 75 mg/kg, 80 mg/kg, 90mg/kg, 100 mg/kg, 125 mg/kg, 150 mg/kg, 175 mg/kg, 200 mg/kg, 225 mg/kg,250 mg/kg, 275 mg/kg, 300 mg/kg, 325 mg/kg, 350 mg/kg, 375 mg/kg, 400mg/kg, 425 mg/kg, 450 mg/kg, 475 mg/kg, 500 mg/kg, 525 mg/kg, 550 mg/kg,575 mg/kg, 600 mg/kg, 625 mg/kg, 650 mg/kg, 675 mg/kg, 700 mg/kg, 725mg/kg, 750 mg/kg, 775 mg/kg, 800 mg/kg, 825 mg/kg, 850 mg/kg, 875 mg/kg,900 mg/kg, 925 mg/kg, 950 mg/kg, 975 mg/kg or 1000 mg/kg, or may rangebetween any two of the foregoing values. In some embodiments, the doseabove may be the total daily dose, or may be the dose administered inone of the one, two or three daily administrations. In some embodiments,the product is administered at a total daily dose of from approximately0.25 g/m² to 4.0 g/m² body surface area, e.g., at least about 0.5, 0.6,0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9 or 2g/m², or up to about 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7,1.8, 1.9, 2.0, 2.2, 2.5, 2.7, 3.0, or 3.5 g/m² or may range between anytwo of the foregoing values. In some embodiments, the product may beadministered at a total daily dose of about 0.5-2.0 g/m² body surfacearea, or 1-1.5 g/m² body surface area, or 0.5-1 g/m² body surface area,or about 0.7-0.8 g/m² body surface area, or about 1.35 g/m² body surfacearea, or about 1.3 to about 1.95 grams/m2/day, or about 0.5 to about 1.5grams/m2/day, or about 0.5 to about 1.0 grams/m2/day, preferably at afrequency of fewer than four times per day, e.g. three, two or one timesper day. Salts or esters of the same active ingredient may vary inmolecular weight depending on the type and weight of the salt or estermoiety. For administration of the dosage form, e.g., a tablet or capsuleor other oral dosage form comprising the enterically coated product, atotal weight in the range of approximately 100 mg to 1000 mg is used. Incertain embodiments, the amount of active ingredient in a tablet orcapsule is approximately 15, 20, 25, 50, 75, 100, 125, 150, 175, 200,250, 300, 400 or 500 mg. Administration may continue for at least 3months, 6 months, 9 months, 1 year, 2 years, or more.

The compositions of the invention can be used in combination with otherdrugs or therapies for each indication contemplated herein. Combinationswith other drugs or therapies that are part of the standard of care foreach indication are specifically contemplated herein.

The compounds and other drugs/therapies can be administered incombination either simultaneously in a single composition or in separatecompositions. Alternatively, the administration is sequential. In someembodiments, the patient is pre-dosed with the compound before theadministration of the other drug/therapy.

The effectiveness of a method or composition of the described herein canbe assessed, for example, by measuring leukocyte cystine concentrationsin subjects affected by cystinosis. Additional measures of the efficacyof the methods of the disclosure include assessing relief of symptomsassociated with fatty liver disease including, but not limited to, liverfibrosis, fat content of liver, incidence of or progression ofcirrhosis, incidence of hepatocellular carcinoma, elevated hepaticaminotransferase levels, increased alanine aminotransferase (ALT),increased aspartate aminotransferase (AST), and elevated serum ferritin.Dosage adjustment and therapy can be made by a medical specialistdepending upon, for example, the severity of fatty liver disease and/orthe concentration of cystine. For example, treatment of fatty liverdisease may result in a reduction in hepatic transaminase of betweenapproximately 10% to 40% compared to levels before treatment. In arelated embodiment, treatment results in a reduction in alanineaminotransferase levels in a treated patient to approximately 30%, 20%or 10% above normal ALT levels, or at normal ALT levels (≥40 iu/L). Inanother embodiment, treatment with cysteamine product results in areduction in aspartate aminotransferase levels in a patient toapproximately 30%, 20% or 10% above normal AST levels or back to normalAST levels.

The methods of the invention also include use of a compound as disclosedherein in preparation of a medicament for treatment of a disease inwhich administration of cysteamine is indicated, and use of a compoundas disclosed herein in preparation of a medicament for administration incombination with another agent for treating a disease whereadministration of cysteamine is indicated. Further provided are kitscomprising a compound as disclosed herein for treatment of a disease inwhich administration of cysteamine is indicated, optionally anotheragent for treatment, and instructions for use in treatment of a diseasewhere administration of cysteamine is indicated.

Animal Models

The compounds disclosed herein can be evaluated in animal models knownin the art for the disease indications contemplated herein.

For example, a number of mouse models which exhibitsteatosis/steatohepatitis exist and include genetically alteredleptin-deficient (ob/ob) or leptin resistant (db/db) and the dietarymethionine/choline deficient (MCD) model. Studies comparing male andfemale rats of varying strains (Wistar, Sprague-Dawley, Long-Evans) witha mouse strain (C57BL/6) as models for NASH can be carried out. Morerecently the use of supra-nutritional diets in animals has resulted in aNAFLD model that physiologically more resembles the human phenotype. Themedical conditions most commonly associated with NAFLD are obesity, TypeII diabetes and dyslipidemia. These conditions can be induced by feedingmice and rats with high fat or sucrose diets. Rats fed with a >70%fat-rich diet for 3 weeks develop pan-lobular steatosis, patchyinflammation, enhanced oxidative stress, and increased plasma insulinconcentrations suggesting insulin resistance. NASH mice have beeninduced through intragastric overfeeding. Mice were fed up to 85% inexcess of their standard intake for 9 weeks. The mice became obese with71% increase in final body weight; they demonstrated increase whiteadipose tissue, hyperglycemia, hyperinsulinemia, hyperleptinemia,glucose intolerance and insulin resistance. Of these mice 46% developedincreased ALT (121=1-27 vs 13+/−1 U/L) as well as histologic featuressuggestive of NASH. The livers of the overfed mice were about twice aslarge expected, beige in color with microscopic evidence of lipiddroplets, cytoplasmic vacuoles and clusters of inflammation.

Mouse models of NASH are created through specific diets (methioninecholine deficient, MCD) or intragastric overfeeding. These mice developserologic and histologic features of NASH. NASH mice are useful inscreening and measuring the effects the compounds disclosed herein onNASH related disease and disorders.

Animal models for kidney fibrosis are known in the art and described,for example, in Eddy et al., “Investigating mechanisms of chronic kidneydisease in mouse models” Pediatr Nephrol. 2011 Jun. 22.

Animal models of Huntington's Disease and Parkinson's disease aredescribed in the art and useful to determine the effects of thecompounds disclosed herein in subjects suffering from disease. See,e.g., Karpuj et al., “Evidence for a role for transglutaminase inHuntington's disease and the potential therapeutic implications.”Neurochem Int. (2002) January; 40(1):31-6, and Bove et al.,“Neurotoxin-based models of Parkinson's disease.” Neuroscience. 2011,Nov. 10.

Additional animal models for other indications are available in the artand are useful to measure the efficacy of the compounds disclosed hereinin said disorders.

While the disclosure has been described in conjunction with specificembodiments thereof, the foregoing description as well as the exampleswhich follow are intended to illustrate and not limit the scope of thedisclosure. Other aspects, advantages and modifications within the scopeof the disclosure will be apparent to those skilled in the art.

EXAMPLES General Methods Cystine Reactivity Assay

Cystine reactivity was assessed by incubating 3 mM test compound with100 nM BODIPY® FL L-Cystine (Life Tech.) at 30° C., pH 7.4 in phosphatebuffer. The initial rate of fluorescence (excitation λ485 nm andemission λ535 nm) increase was used as a measure of cystine reactivityand hence cysteine formation. Compound cystine reactivity was expressedas a percentage of that of 3 mM cysteamine.

Under resting conditions, the inherent fluorescence of BODIPY® FLL-cystine was very low due to the spatial proximity of the two BODIPYmolecules causing quenching. Under chemical reaction (disruption of theS—S bond), the spatial distance was increased, the quenching lost, andmeasured fluorescence increased. Time (minutes) vs. fluorescence(relative fluorescence units—RFU) was plotted and linear regressionanalysis was performed to generate a slope value corresponding to thereaction rate in RFU/min and an R2 value.

ADO Metabolism Assay

The rate of ADO metabolism was determined by incubating 15 mM testcompound with 60 μg/ml human recombinant ADO and 20 nM MitoXpress®02-sensitive fluorescent probe (Luxcel) at 37° C. in assay buffer/saltin a sealed assay chamber. The initial rate of fluorescence (excitationλ380 nm and emission λ650 nm) increase was used to measure ADO-dependentoxygen depletion activity. The ability of ADO to metabolize compoundsand concomitantly consume oxygen was expressed relative to that of 15 mMcysteamine.

Cystine Depletion Assay

The ability of test compounds to deplete cystine from cystinotic humanfibroblasts (Coriell) was determined by incubating compounds with cellsfor 60 min at 37° C., 95% (v/v) air/5% (v/v) CO₂ in Minimal EssentialMedia Eagle. Treated cell samples were subsequently harvested, thenhomogenized to ensure plasma and lysosomal membrane disruption. Proteinwas removed by acid precipitation and cystine in the supernatantmeasured using LC-MS/MS and HILIC chromatography. Protein levels weredetermined and cystine levels reported as nM per mg/ml protein. Cystinelevels after test compound treatment were expressed as a percentage ofthat of untreated control cells.

Rat Hepatocyte Assay

Rat hepatocytes were used to model in vivo hepatic clearance of thecompounds by the ADO enzyme. Briefly, a Hepatocyte Stability Assay wascarried out using cryopreserved rat hepatocytes. The test compound wasadded to cells (e.g., 10⁶ cells) at a concentration of approximately 3μM (50 μL of 10 mM solution). Other concentrations can also be used.Presence of compound was measured at varying timepoints (0, 5, 15, 30,45, 60, 90, 120 minutes) by LC-MS/MS analysis and intrinsic clearance(CL_(int)) rate was measured. See, e.g., Lubberstedt et al., HepaRGhuman hepatic cell line utility as a surrogate for primary humanhepatocytes in drug metabolism assessment in vitro, J Pharmacol ToxicolMethods 63:59-68, 2010; or Zanelli et al., Comparison of cryopreservedHepaRG cells with cryopreserved human hepatocytes for prediction ofclearance for 26 drugs, Drug Metab Dispos 40:104-110, 2012.

Rodent Pharmacokinetic Assays

Pharmacokinetic parameters of a test compound in a rodent species (mouseand/or rat) were determined by administering a test compound at 2 and 10mg free base equivalents per kg via the intravenous and oral gavageroute to groups of three animals per route.

Blood samples were taken at various times after administration, plasmaprepared, and submitted to analysis for parent drug using a qualifiedLC-MS-MS assay. Pharmacokinetics parameters derived from the plasmaanalytical data were determined using non-compartmental analysis.

Example 1 Activity of N-Substituted Compounds

Compounds were assayed at a concentration of 50 μM or 100 μM accordingto the general methods described above.

TABLE 1 Compound Cysteamine 1a 1b 1c Structure

Cystine 6543 — 7933 11027 Reactivity ADO 111%  4%   9%  2% metabolismCystine 101% 85% 104% 98% depletion (106%) (94%) (104%) Rat Hepatocytes33.7 — 27.0 26.3 CL _(int) (ml/min/mg) Rat PK -Half 0.5 h — 0.3 h 0.3 hlife 62% —  15% 28% Bioavailability Mouse PK -Half 2.4 h — — — life Bio- 33% availability ( ) = disulfide efficacy

As shown in Table 1, compounds 1b and 1c demonstrate increasedreactivity with cystine compared to cysteamine, along with similarcystine depletion levels. Compound 1a demonstrates slightly decreasedcystine depletion levels as compared to the cystine depletion levels forcysteamine. Advantageously, compounds 1a, 1b, and 1c demonstratesignificantly reduced metabolism by ADO compared to cysteamine.

Example 2 Activity of Cyclic N-Substituted Compounds

Compounds were assayed at a concentration of 50 μM or 100 μM accordingto the general methods described above.

TABLE 2 Compound Cysteamine 2a 2b 2c 2d Structure

Cystine 6543 — 12392 — 7332 Reactivity ADO 111%  7%  4%  2%    1%metabolism Cystine 101% 82% 99% 73%   79% depletion (106%) Rat 33.7 —15.7 —   12.6 Hepatocytes CL _(int) (ml/min/mg) Rat PK -Half 0.5 h — — —2.3 h life Bio-  62%   34% availability Mouse PK -Half 2.4 h — 0.4 h 7.1h 3.9 h life Bio-  33% 33% 74%   38% availability ( ) = disulfideefficacy

As shown in Table 2, compounds 2b and 2d demonstrate increasedreactivity with cystine compared to cysteamine. The compounds alsodemonstrates similar (compound 2b) or slightly decreased (compounds 2a,2c, 2d) cystine depletion levels as compared to the cystine depletionlevels for cysteamine. Advantageously, compounds 2a, 2b, 2c, and 2ddemonstrate significantly reduced metabolism by ADO compared tocysteamine.

Example 3 Activity of Alkyl Chain-Substituted Compounds

Compounds were assayed at a concentration of 50 μM or 100 μM accordingto the general methods described above.

TABLE 3 Compound Cysteamine 3a 3b 3c 3d Structure

Cystine 6543 2284 — 7733 — Reactivity ADO 111% 14% —  9% 14% metabolismCystine 101% 32% 97% 97% 55% depletion (106%) Rat 33.7 — — 29.5 —Hepatocytes CL _(int) (ml/min/mg) Rat 0.5 h — — 0.8 h — PK -Half  62%23% life Bio- availability Mouse 2.4 h — — — — PK -Half  33% life Bio-availability ( ) = disulfide efficacy

As shown in Table 3, compound 3c demonstrates increased reactivity withcystine compared to cysteamine. The compounds also demonstrate similar(compounds 3b, 3c) or slightly decreased (compounds 3a, 3d) cystinedepletion levels as compared to the cystine depletion levels forcysteamine. Advantageously, compounds 3a, 3c, and 3d demonstratesignificantly reduced metabolism by ADO compared to cysteamine.

Example 4 Sulfur-Nitrogen Distances and Activity of Compounds

The sulfur-nitrogen distances for each of the compounds in Table 4 werecalculated using quantum mechanic calculations according to thefollowing procedure. First, the 2D chemical structures were read intoSpartan '14 and automatically converted to 3D models. Each 3D structurewas then minimized using MMFF (Merck Molecular Force Field) (compound inneutral form). Then, to each minimized MMFF structure (neutral form),quantum mechanics calculations were run as follows: EquilibriumGeometry; Density Functional, B3LYP, 6-31G*; vacuum; in neutral form; nounpaired electrons.

Cystine depletion levels for the compounds are expressed in Table 4 as apercent relative to the cystine depletion achieved by cysteamine (c.f.cysteamine 100%) at a similar concentration.

Vortex® scatter plots (generated from Dotmatics®) were used tographically represent the relationship between the sulfur-nitrogendistance and the extent of cystine depletion for the compounds. As shownin FIG. 1, high levels of cystine depletion are observed for compoundshaving a sulfur-nitrogen distance of about 3.6 to 4.7 Å, and inparticular, about 4.1 Å.

Unless indicated (by the term “Abs”), relative stereochemistry is shownin Table 4.

TABLE 4 Cystine Cystine Reactivity Reactivity Assay Assay ADO (30° C.)(RT) Metabolism S-N Cystine (%) (RFU) Assay (%) Cmpd. Dist. DepletionMean Mean Mean Compound No. (Å) (%) Value Value Value

42694 4.2 20 <50 — —

42693 3.1 110 >90 — —

42692 3.1 100 >90 — —

42691 5.3 — 50-90 — —

42690 4.1 — — — —

42689 4.1 — — — —

42618 4.7 49 — — —

42617 4 101 50-90 — <25

42616 4 79 50-90 — <25

42615 4.7 106 <50 — <25

42614 4.7 117 — — <25

42613 4.1 109 >90 — <25

42612 4.2 98 — — —

42611 5.9 55 >90 — <25

42610 3.1 91 >90 — —

42609 4.7 17 — — —

42494 4.6 88 50-90 — <25

42493 4.1 74 50-90 — <25

42492 6.2 — <50 — —

42491 5.1 — <50 — <25

42460 4.7 111 50-90 — <25

42458 4.7 91 >90 — —

42457 3.8 108 50-90 — <25

42391 4.7 60 >90 — —

42389 4.5 36 50-90 — <25

42388 4.1 97 >90 — <25

42345 3.4 50 >90 — <25

42344 4.7 38 — — —

42343 3.8 84 — — —

42342 3.8 91 — — —

42341 4.7 81 <50 — <25

42338 4.6 — <50 — —

42248 4.1 103 >90 — <25

42247 4.1 96 >90 — <25

42213 3.7 0 >90 — >75

42212 3.3 — <50 — —

42211 4.1 79 >90 — <25

42210 4.1 87 >90 — <25

42209 4.8 99 50-90 — <25

42208 4.1 102 >90 — <25

42153 4.6 — — — —

42152 4.1 126 — — —

42151 3.8 133 — — —

42099 4.1 79 — — 25-75

42098 4.1 70 — — <25

42097 4.1 97 — — —

41448 3.1 55 — — 25-75

41446 3.7 82 — — <25

41188 4.7 87 50-90 — <25

18781 4.1 92 — — <25

17522 6.9 6 — <4000 <25

17521 4.7 79 50-90 4000-9000 <25

17518 4.1 99 — >9000 <25

17517 4.1 98 — >9000 <25

17515 4.1 97 — 4000-9000 <25

17333 4.1 104 — 4000-9000 <25

17330 3.2 — — 4000-9000 25-75

17329 5.2 36 — <4000 <25

17328 4.1 13 — <4000 <25

17287 4.1 — — <4000 <25

1585 4.1 100 >90 4000-9000 >75

18782 4.1 32 — <4000 <25

Example 5 Activity of Compounds

Cystine depletion levels for each of the compounds in Table 5 areexpressed as a percent relative to the cystine depletion achieved bycysteamine (c.f. cysteamine 100%) at a similar concentration.

Unless indicated (by the term “Abs”), relative stereochemistry is shownin Table 5.

TABLE 5 Cystine ADO Cystine Reactivity Assay Metabolism Cmpd. Depletion(%) (30° C.) (%) Assay (%) Compound No. Mean Value Mean Value Mean Value

152071 70 — —

151272 11 — —

151271 27 — —

151270 3 — —

151182 59 50-90 —

151181 69 50-90 —

150731 58 >90 —

150730 14 >90 <25

149611 27 >90 —

149610 15 <50 <25

149609 50 <50 <25

149608 55 <50 <25

149607 17 >90 —

145696 32 >90 <25

145695 0 >90 <25

145694 0 >90 <25

145693 81 50-90 <25

145692 75 — —

145691 0 — —

145611 9 <50 <25

145610 11 <50 <25

145609 121 — <25

145592 156 >90 —

145591 61 50-90 <25

145590 104 >90 <25

145589 70 50-90 <25

145588 31 >90 <25

145587 13 >90 <25

145586 21 50-90 <25

145585 11 >90 —

145584 0 >90 <25

145566 16 <50 <25

145565 31 — —

145564 36 >90 25-75

145232 16 <50 <25

42460 78 50-90 —

42457 77 50-90 —

42388 63 >90 <25

42341 83 <50 —

42210 89 >90 —

41446 82 >90 <25

2415 14 — —

2403 19 <50 <25

1176 10 <50 <25

1141 0 <50 —

981 25 <50 <25

653 23 <50 <25

Example 6 Dimethyl Sulfide (DMS) Detection in Rat Whole Blood FollowingOral Compound Administration

Male Sprague Dawley (SD) rats (200-250 g) were housed in cages under a12 hour light/dark cycle with free access to food and water. Temperatureand humidity were controlled according to UK Home Office regulations.For oral (po) administration, test compounds were formulated in water ata concentration of 30 mg/mL, to provide a dose of 150 mg/kg whenadministered orally in a 5 mL/kg dosing volume. Following compound orwater (vehicle) administration to rats (n=15 per compound), terminalblood samples (>5 mL) were taken by cardiac puncture under CO₂ atdefined time-points post oral dosing (pre-dose, 0.25 hour, 0.5 hour, 1hour and 2 hour). Blood was placed in sealed heparinized Eppendorf tubeson wet ice and stored at 4° C. prior to analyzing DMS levels by gaschromatography (GC).

For each sample of rat blood, 1 mL was added to an evacuated 20 mLGC-Headspace vial containing tetrahydrofuran (THF) internal standard.Headspace vials were sealed with a crimp cap and septum (to prevent thevaporization of volatile sulfur compounds into the open air) andthoroughly vortex mixed. All samples were processed using gaschromatography to measure DMS levels. In short, sulfur containingcompounds were thermally liberated into the gas carrier stream andinjected (1 μL) into the gas chromatography column (30 m×0.32 mm DB-6241.8 μm). Under these conditions, the retention time for dimethyl sulfide(DMS) and THF internal standard were approximately 1.83 minutes and 3.82minutes respectively.

For each analyzed blood sample, the ratio of DMS peak area to THF peakarea was calculated. For the DMS calibration curve and assay linearitychecks, this ratio was graphically plotted against the concentration ofreference DMS standards (blank, 60 nM, 180 nM, 360 nM, 600 nM, 1200 nM,1800 nM and 3000 nM). The concentration of DMS in each blood sample wascalculated from the equation produced by this plot by linear regressionanalysis. The results are shown in Table 6. The lower limit of detectionwas 60 nM and a value of <60 indicates no DMS peak was detected in thesample chromatogram. Advantageously, the tested compounds producedreduced levels of DMS compared to cysteamine, suggesting that thesecompounds have reduced unpleasant side effects (e.g., halitosis)compared to cysteamine therapy.

TABLE 6 Mean DMS levels (nM) ± STD Cmpd. Dose Pre- Time Time Time TimeCompound No. (mg/kg) po Dose 0.25 hr 0.5 hr 1 hr 2 hr

1585 150 <60 641 ± 324 1097 ± 418 1162 ± 373 1437 ± 334

42341 150 <60 <60 <60 <60 <60

42210 150 <60 <60 No data <60 <60

41188 150 <60 <60 No data <60 <60

41446 150 <60 <60 No data <60 <60

18782 150 <60 <60 No data <60 <60

Example 7 Glutamate-Induced Neuronal Excitotoxicity Assay

The ability of test compounds to inhibit glutamate-inducedexcitotoxicity (neuroprotection) in St-HdhQ^(111/111) cells wasdetermined by incubating compounds with cells for 60 min at 33° C., 95%(v:v) air/5% (v:v) CO₂ in Dulbecco's Modified Eagle Media. Followingthis, excitotoxicity was induced by the addition of 6 mM L-glutamate for24 hours. Cell viability was assessed by measuring ATP levels using aluminescent-based CellTitre Glo assay (Promega). Compoundneuroprotection (% cell survival) was expressed as a % of the effectrecorded with 100 μM cysteamine (denoted as 100% cell survival).Advantageously, the tested compounds resulted in high levels of cellsurvival (e.g., at least 50% cell survival when expressed as a % of theeffect for 100 μM cysteamine), including levels of cell survival highlysimilar to that of cysteamine (e.g., at least 80% cell survival whenexpressed as a % of the effect for 100 μM cysteamine), suggesting thatthese compounds have similar neuroprotection levels as compared tocysteamine therapy.

TABLE 7 HdhQ^(111/111) cell % survival (mean ± STD) Compound/ Test Conc.Test Conc. Compound No. 100 μM 10 μM 151182 85 ± 11 1 ± 5 151181 90 ± 9 3 ± 8 149611 102 ± 13  35 ± 13 149607 98 ± 12 32 ± 9  145695 74 ± 10  8± 10 145611 94 ± 8  24 ± 13 145610 87 ± 10 11 ± 8  145592 88 ± 8  17 ±10 145590 90 ± 11 14 ± 7  145588 98 ± 5  9 ± 9 145587 97 ± 7  52 ± 9 145586 89 ± 9  9 ± 6 145585 100 ± 9   13 ± 10

86 ± 10 3 ± 7

93 ± 9  13 ± 13

95 ± 6  11 ± 11  42693 77 ± 9  80 ± 10  42692 71 ± 5  82 ± 4   42617 89± 7   7 ± 10  42616 56 ± 7  5 ± 7  42614 59 ± 12 2 ± 6  42610 77 ± 8  19± 6   42493 87 ± 10  6 ± 10  42460 95 ± 9  3 ± 5  42457 91 ± 6  6 ± 5 42388 91 ± 5  5 ± 6  42341 55 ± 21  1 ± 10  42248 81 ± 6  2 ± 7  4224784 ± 13 5 ± 8  42211 84 ± 19 2 ± 9  42210 105 ± 8   7 ± 6  41446 101 ±11  6 ± 9  41188 105 ± 11  20 ± 6    1585 105 ± 13  20 ± 6  145694 15 ±15  2 ± 10

16 ± 7  1 ± 6  42494 28 ± 17 0 ± 7

Numerous modifications and variations in the invention as set forth inthe above illustrative examples are expected to occur to those skilledin the art. Consequently only such limitations as appear in the appendedclaims should be placed on the invention.

What is claimed:
 1. A compound having the following structure or adisulfide thereof:

wherein R¹ is C₁₋₅alkyl; and R³, R⁴, R⁵, R⁷, and R⁸ are independentlyselected from the group consisting of H and C₁₋₅ alkyl.
 2. A compoundhaving the following structure or a disulfide thereof:

wherein R¹, R³, R⁴, R⁵, R⁶, and R⁷ are independently selected from thegroup consisting of H and C₁₋₅alkyl.